Changes of receptor mRNAs for oxytocin and estrogen during the estrous cycle in rat uterus

Oxytocin receptor (OTR) mRNA levels increase dramatically near term and is potently stimulated by estrogen because increased OTR mRNA levels result from estrogen treatment in ovariectomized rat uterus. In this study, OTR, estrogen receptor (ER) alpha and ERbeta mRNA levels in the rat uterus during the estrous cycle were examined by quantitative RT-PCR. OTR mRNA levels during the estrous cycle began to increase on diestrus (P<0.05, vs value on estrus), reached maximal increase both in the morning (1000-1130 hr) and afternoon (1600-1630 hr) on proestrus (P<0.01, vs metestrus, diestrus and estrus) and then declined on estrus. In contrast ER alpha mRNA levels began to decrease on diestrus, reached statistical significance both in the morning and the afternoon on proestrus (P<0.01, vs metestrus, diestrus and estrus) and returned to the value of metestrus on estrus. ERbeta mRNA levels were low in the morning and the afternoon on proestrus (P<0.01, vs metestrus and estrus) and also returned to metestrus values on estrus. Treatments with estrogen for 3 days significantly decreased both ERalpha and ERbeta mRNA levels. It can be concluded from these results that during the estrous cycle, OTR mRNA levels in rat uterus predominantly increase at proestrus with a decrease in ERalpha and ERbeta mRNA levels, which is probably due to the increased estrogen levels in circulation before ovulation.     

Expression of Frizzled 2 in the mouse ovary during oestrous cycle

The Wnt/β‐catenin pathway is a conserved signalling pathway that regulates gene expression and controls diverse developmental processes such as cell fate specification, cell proliferation and cell differentiation. To our knowledge, the potential role of this pathway in the adult ovary has been poorly addressed. To this end, we investigated the expression pattern of Frizzled 2 in the mouse ovary during oestrous cycle by real‐time Q‐PCR, in situ hybridization, Western blotting and immunohistochemistry. In this study, we used uterine wet weight and H3.2 mRNA level as two markers for classification of mouse oestrous cycle. We found that both uterine wet weight and H3.2 mRNA are the highest in the proestrus and decrease from oestrus to diestrus. During oestrous cycle, Frizzled 2 mRNA and protein exhibited the highest level in the proestrus stage and rapidly reduced from oestrus to diestrus. In situ hybridization results showed that the positive signals for Frizzled 2 are highly detected in the oocyte and stroma in proestrus stage, while moderate or weak Frizzled 2 mRNAs are localized in the oocyte, granulosa cells, stroma and corpus luteum from oestrus to diestrus. The localization pattern of Frizzled 2 protein is mostly consistent with its mRNA, but stronger Frizzled 2 proteins are present in the granulosa cells and membrane of oocyte in oestrous cycle. Our data suggest that Frizzled 2 may be involved in regulating follicle growth, oocyte maturation and luteinization during oestrous cycle.     

Effects of natural and synthetic PGF2 alpha preparations on the uterine contractility of cattle

The effects of luteolytic doses of PGF2 alpha (25 mg Dinoprost) and its synthetic analogues Cloprostenol (500 micrograms), Luprostiol (15 mg) and Tiaprost (525 micrograms) on bovine myometrial activity were investigated using a miniature pressure transducer placed in one uterine horn. The compounds were administered intravenously to 4 lactating cyclic cows at diestrus, proestrus, estrus and metestrus. Intrauterine pressure changes were assessed by computerized planimetry of the pressure tracings 30 minutes before and 60 minutes after treatment. Baseline intrauterine pressure was set at zero and treatment effects were expressed as percent change from an equivalent control period (= 100%). Following administration of Dinoprost there was a significant increase of uterine contractility in diestrus (515%), proestrus (198%) and metestrus (256%), but not in estrus. In comparison to PGF2 alpha the analogues Luprostiol and Tiaprost were less effective (Luprostiol: 195% and 154% in diestrus and proestrus resp., Tiaprost: 215% during diestrus), while Cloprostenol did not cause a significant change of intrauterine pressure in any stage of the estrous cycle. The results indicate that the myotonic effects which F2 alpha-prostaglandins exert on the uterus of cycling cows is affected both by the type of prostaglandin and the stage of the estrous cycle.     

Uterine motility during the estrous cycle: studies in healthy bitches

Natural and drug-induced uterine motilities (UM) were recorded via uterine-implanted catheter-tip pressure transducers in 5 bitches during proestrus and estrus and in 6 bitches during early (30 days) and late (60 days) diestrus. The UM was monitored in unanesthetized bitches for 2 days after the estrous-cycle stage had been determined by vaginal cytologic findings. Natural UM, expressed as Alexandria units (pressure X frequency X duration of contractions), was greatest during estrus, moderate during proestrus, and appreciably decreased or lacking during early and late diestrus. During proestrus and estrus, prostaglandin F2 alpha (5 micrograms/kg of body weight, IV) induced intrauterine pressures of 107 and 115 mm of Hg, respectively, and oxytocin (0.05 USP units/kg, IV) induced pressures of 106 and 116 mm of Hg, respectively. In contrast, the intrauterine pressure values induced by prostaglandin F2 alpha given IV during early and late diestrus were 61 and 58 mm of Hg, respectively, and for oxytocin, were 61 and 51 mm of Hg, respectively. Prostaglandin F2 alpha given IM (50 micrograms/kg) also induced a greater intrauterine pressure during proestrus and estrus than during diestrus. We concluded that in the bitch, natural and drug-induced UM are decreased during early and late diestrus.     

Efficacy and toxicity of estrogens commonly used to terminate canine pregnancy

Three groups of bitches were treated with diethylstilbestrol (75 micrograms/kg) orally for 7 days (n = 12), estradiol cypionate intramuscularly once (22 micrograms/kg; n = 12), or estradiol cypionate intramuscularly once (44 micrograms/kg; n = 12). Treatments commenced during late proestrus (n = 4/group), the fourth day of behavioral estrus (n = 4/group), or the second day of diestrus (n = 4/group). All bitches were bred on alternate days throughout estrus to stud dogs of known fertility. Ovariohysterectomies were performed on day 25 of diestrus to diagnose pregnancy and to assess any pathologic changes in the uterus. Eleven bitches treated with diethylstilbestrol, 6 bitches treated with the low dosage of estradiol cypionate, and 4 bitches receiving the high dosage of estradiol cypionate were pregnant at the time of surgery. Ten of the bitches treated with estrogens during proestrus, 6 treated during estrus, and 4 treated during diestrus were pregnant. The serum concentration of progesterone in 2 bitches treated with the high dosage of estradiol cypionate decreased to less than 2 ng/ml by day 25 of diestrus, suggesting premature luteal regression. Diethylstilbestrol appeared to have little efficacy in terminating pregnancy. Estradiol cypionate appeared to have greater efficacy when administered during estrus or early diestrus; however, pyometra developed in 2 bitches treated with this estrogen during diestrus.     

发情周期不同阶段牦牛子宫中黄体生成素受体的表达变化

为研究发情周期不同阶段牦牛子宫中黄体生成索受体(LHR)的定位及表达变化,笔者利用免疫组织化学SP法分别检测发情期、发情后期、间情期和发情前期牦牛子宫中LHR的表达,并进行光密度值分析.结果表明,LHR免疫阳性产物在牦牛子宫腺上皮细胞、基质细胞、血管内皮细胞、血管平滑肌细胞和肌层平滑肌细胞中均有表达;腺上皮细胞、基质细胞和肌层平滑肌细胞中LHR在发情前期和发情期表达最弱,发情后期表达增加,间情期表达最强(P＜0.05);子宫内膜血管平滑肌细胞中LHR的表达在发情期最强,间情期最弱(P＜0.05);血管内皮中LHR在发情期和发情前期表达很强,发情后期和间情期显著下降(P＜0.05).结果表明LHR参与了发情周期不同阶段牦牛子宫功能变化的调控.     

Recovery and light microscopic evaluation of follicular oocytes of swine and relationship between the degeneration rate of oocytes and the estrus phase

Cumulus-oocyte complexes were recovered from 25 gilts by aspiration of follicular fluid or cutting of follicles from all Graafian follicles of greater than or equal to 3 mm in diameter during diestrus, proestrus or estrus. In 5 gilts the oocytes were collected post ovulation by flushing of oviducts. The recovery rate of follicular oocytes differed between 75.5% during the late diestrus (days 13-17) and 43.5% during the proestrus (days 18-21). During the proestrus and on day 1 of the estrus the recovery of oocytes was more difficult as a result of the higher viscosity of follicular fluid and the mucification of cumulus-oocyte complexes. The degeneration rate of oocytes was high during the diestrus with a peak at the time of regression of corpora lutea. From diestrus to the estrus the degeneration rate decreased. Following degeneration rates were found in the oocytes during the cycle: days 7-12: 38.8%, days 13-17: 50.0%, days 18-21: 29.6%, day 1 of the estrus: 10.8%, day 2 of the estrus ante ovulation: 11.8%, day 2 of the estrus post ovulation: 6.2%. Signs of degeneration were: Loss of cumulus cells (during diestrus and proestrus), damaged zona pellucida, enlargement of perivitelline space, deformation of oocyte, alteration of structure of the ooplasm, diameter of vitellus less than 100 microns. It was concluded that the selection of dominant follicles takes place in pigs during a long time of the cycle, especially during the diestrus. There were not any indications of a 2-wave hypothesis of follicular growth during the cycle in pig.     

Concentration of receptors for estradiol and progesterone in canine endometrium during estrus and diestrus

Receptors for estrogen and progesterone were measured in cytosols prepared from specimens of canine endometrium obtained at late proestrus, day 4 of estrus, day 2 of diestrus, and at 10 day intervals from days 10 through 80 of diestrus. Twenty nine adult bitches were used, with 2 to 4 dogs used at each time point. Concentrations of estradiol receptors measured in endometrial cytosols from late proestrus through day 10 of diestrus were similar (mean +/- SEM: 9.9 +/- 2.2, 10.5 +/- 1.2, 16.3 +/- 1.6, and 16.2 +/- 2.9 pmol/g of tissue at proestrus, day 4 of estrus, days 2 and 10 of diestrus, respectively). As serum concentrations of progesterone increased during early diestrus, the concentration of estradiol receptors decreased and were significantly (P less than 0.05) lower on days 30 (4.9 +/- 1.3 pmol/g of tissue) and 40 (3.7 +/- 0.6 pmol/g of tissue) of diestrus. After day 40 of diestrus, when serum concentrations of progesterone were approaching basal concentrations, the concentration of estradiol receptors increased and remained significantly (P less than 0.05) higher from days 60 to 80 of diestrus (day 60, 13.4 +/- 2.9; day 70, 15.7 +/- 1.7; day 80, 19.8 +/- 2.4 pmol/g of tissue). As observed for estrogen receptors, the concentration of endometrial receptors for progesterone also gradually increased from late proestrus (4.9 +/- 1.3 pmol/g of tissue) to day 2 of diestrus (6.4 +/- 0.3 pmol/g of tissue).(ABSTRACT TRUNCATED AT 250 WORDS)     

Expression and regulation of Foxa2 in the rat uterus during early pregnancy

The forkhead box a (Foxa) protein family has been found to play important roles in mammals. Recently, the expression of Foxa2 was reported in the mouse uterus, and it was reported to be involved in regulation of implantation. However, the regulation of Foxa2 expression in the uterus is still poorly understood. Therefore, the present study was conducted to investigate the expressional profiles of Foxa2 in the rat uterus during the estrus cycle and pregnancy. Furthermore, the effect of steroid hormones and Hedgehog protein on the expression of Foxa2 was analyzed in vivo and in vitro. In this study, the level of expression of Foxa2 was low in the rat uterus during the different stages of the estrus cycle. However, the expression increased transiently during early pregnancy at 3.5 days post coitus (dpc) and decreased at 5.5 dpc. In ovariectomized rats, P4 treatment had no effect on the expression of Foxa2 compared with the expression in control animals. Moreover, the expression of Foxa2 in cultured epithelial cells was not increased by P4 treatment in vitro. However, Foxa2 expression was significantly decreased in the rat uterus after 24 h of E2 treatment. Treatment of cells with a recombinant Hedgehog protein significantly increased the expression of Foxa2. These results suggest that the expression of Foxa2 may transiently increase just before the implantation and it may be regulated by E2 and Hedgehog protein.     

Chemotactic properties and protein of equine uterine fluid

Forty uterine fluid samples were obtained from 4 mares classified as resistant to uterine bacterial infection. The uterus of each mare was flushed with 50 ml of saline solution during estrus and diestrus of successive estrous cycles. Bacteria or fungi were isolated from 4 samples, and 7 additional samples were obtained from a mare with active intrauterine infection. Fluid volumes obtained during estrus (means = 40.3 +/- 11 ml) tended to be greater than those recovered during diestrus (means = 36.8 +/- 7.9 ml), but the difference was not significant. Concentrations and yields of protein in recovered fluid did not vary significantly with the day of the estrous cycle. Protein concentration was significantly increased in samples from the infected uterus (P = less than 0.001). The ability of uterine fluid samples to attract neutrophils was measured using chemotactic chambers. There was no significant difference between distances migrated by neutrophils toward fluids obtained during estrus or diestrus. Chemotaxis scores tended to be higher with samples from the infected uterus, and the difference was significant for samples from 2 mares. Chemotaxis was neither significantly correlated with protein concentration of uterine fluid, nor with serum estrogens or progesterone.     

Effect of Conceptus on Transforming Growth Factor (TGF) β1 mRNA Expression and Protein Concentration in the Porcine Endometrium—In Vivo and In Vitro Studies

Transforming growth factor (TGF) β and its receptors are expressed at the conceptus-maternal interface during early pregnancy in the pig. The present studies were conducted to examine: (1) the effect of conceptus products on TGFβ1 mRNA expression and protein concentration in the porcine endometrium using in vivo and in vitro models, and (2) the effect of TGFβ1 on proliferation of porcine trophoblast cells in vitro. During in vivo experiments, gilts with one surgically detached uterine horn were slaughtered on days 11 or 14 of the estrous cycle and pregnancy. For in vitro studies, endometrial explants and luminal epithelial (LE) cells co-cultured with stromal (ST) cells were treated with conceptus-exposed medium (CEM). Moreover, porcine trophoblast cells were treated with TGFβ1, and the number of viable cells was measured. On day 11, the presence of conceptuses had no effect on TGFβ1 mRNA expression, but decreased the TGFβ1 protein concentration in the connected uterine horn compared with the detached uterine horn. In contrast to day 11, on day 14 after estrus, TGFβ1 mRNA expression and protein content in the endometrium collected from the gravid uterine horn were greater when compared with the contralateral uterine horn. The treatment of endometrial slices with CEM resulted in greater TGFβ1 mRNA expression and protein secretion. LE cells responded to CEM with an increased TGFβ1 mRNA level. Moreover, TGFβ1 stimulated the proliferation of day 14 trophoblast cells. In summary, porcine conceptuses may regulate TGFβ1 synthesis in the endometrium at the time of implantation. TGFβ1, in turn, may promote conceptus development by increasing the proliferation of trophoblast cells.     

Studies on Some Aspects of Neutrophil Functions and Uterine Defenses in Cows during the Estrous Cycle

Some aspects of the bovine uterine defense mechanisms were evaluated over two successive estrous cycles in five adult nonpregnant Friesian cows. Uterine flushings and blood samples were taken from each cow during the estrus and diestrus phases of each estrous cycle. A significant higher percentage of Candida albicans (C. albicans) were phagocytosed (P < 0.01) and killed (P < 0.05) by blood neutrophils during estrus than during diestrus. N-acetyl-β-D-glucosaminidase activity of uterine flushings, blood neutrophil alkaline phosphatase activity scores, and serum progesterone concentrations during diestrus increased significantly (P < 0.01) compared to those at estrus. Mean total blood leucocyte and monocyte counts increased significantly (P < 0.05) during estrus compared to diestrus .
It is concluded, that the different hormonal status has at each stage of the estrous cycle a definite effect on uterine resistance against bacterial infection and a fall in serum progesterone concentrations is associated with an increased ability of blood neutrophils to phagocytose and kill C. albicans .     

Expression of interleukin-18 receptor mRNA in the mouse endometrium

Interleukin-18 (IL-18) is a proinflammatory cytokine involved in chronic inflammation, autoimmune diseases, and a variety of cancers, and is expressed in mouse uteri. Our previous study suggested that IL-18 acts as a paracrine factor, regulating endometrial function. To elucidate the physiological roles of IL-18 in the mouse endometrium, the expression of the IL-18 receptor (IL-18R) alpha subunit was analyzed. IL-18Ralpha mRNA was expressed in several mouse organs in addition to the endometrium. In situ hybridization analysis using a biotin-labeled mouse IL-18Ralpha riboprobe demonstrated that IL-18Ralpha mRNA expression was detected in glandular epithelial cells, stromal cells around uterine glands, and myometrial cells in the mouse uterus, suggesting that these cells are targets for IL-18. The uterine IL-18Ralpha mRNA expression level changed with the estrous cycle. The uterine IL-18Ralpha mRNA levels of estrous mice were higher than those of diestrous mice. In addition, the IL-18Ralpha mRNA levels in uteri at 3 and 14 days after ovariectomy were higher than those at diestrus and decreased following treatment with estradiol-17beta or progesterone. These findings suggest that IL-18Ralpha gene expression is regulated by estrogen and progesterone and that the uterine IL-18 system is involved in the regulation of uterine functions in a paracrine manner.     

Tumor necrosis factor-alpha as a possible auto-/paracrine factor affecting estrous cycle in the cat uterus

Tumor Necrosis Factor-alpha (TNFalpha) is a pleiotrophic cytokine, affects either normal or tumor cells, and influences cellular differentiation. TNFalpha role in female reproduction has been proven to be mediated through an influence on prostaglandin (PGs) synthesis and output. To evaluate the possible role of TNFalpha in an auto-/paracrine regulation in the cat uterus, mRNA expression coding for TNFalpha and its receptors (TNFR1 and TNFR2), and TNFalpha protein content at different stages of the estrous cycle were investigated. Additionally, TNFalpha involvement in PG secretion at different stages of the estrous cycle was investigated by in vitro tissue culture. Gene expressions coding for TNFalpha and TNFR1 were the highest at diestrus (P < 0.05). TNFalpha protein expression was the lowest at interestrus (P < 0.05). Nevertheless, TNFR2 was not affected by the estrous stage. TNFalpha at a dose of 1 ng/ml significantly increased PGF2alpha secretion at estrus (P < 0.01) and PGE2 secretion at diestrus (P < 0.001) after 12h incubation. Overall findings indicate that TNFalpha locally produced in the cat's uterus, stimulates PG secretion in an estrous cycle-related manner.     

生殖周期小鼠子宫内膜上皮淋巴细胞的数量变化

应用甲苯胺蓝染色方法，观察了发情周期、妊娠期和分娩后小鼠子宫粘膜上皮内淋巴细胞的消长情况。与间情期相比，上皮内淋巴细胞的数量在发情前期减少，而发情期和发情后期又增加。妊娠 １ ｄ 数量较少，１４ ｄ 开始回升，１８ ｄ 的数量超过间情期。分娩后 １ ｄ 突然升高，随后又逐渐恢复。这些结果提示，上皮内淋巴细胞可能参与子宫粘膜免疫。     

The Activity and Localization of 3β-hydroxysteroid Dehydrogenase/Δ5-Δ4 Isomerase and Release of Androstenedione and Progesterone by Uterine Tissues During Early Pregnancy and the Estrous Cycle in Pigs

Steroid hormones are produced by the porcine uterus. We hypothesized that the uterus in pigs possesses active 3β-hydroxysteroid dehydrogenase/Δ⁵-Δ⁴ isomerase (3β-HSD) responsible for progesterone and androstenedione production, that uterine steroids may supplement the amount of steroid hormones produced by embryos and corpus luteum and that these steroids are necessary for maintenance of pregnancy. In this study, we examined 1) endometrial and myometrial expression of 3β-HSD mRNA, 2) uterine 3β-HSD protein activity and 3) in vitro production of A₄ and P₄ by uterine slices harvested from pigs on days 10 to 11, 12 to 13 and 15 to 16 of pregnancy and the estrous cycle. The expression of 3β-HSD and the presence and activity of 3β-HSD protein were different in the endometrium and the myometrium during the examined periods of pregnancy and the estrous cycle. Production of A₄ by the endometrium and myometrium was highest on days 12 to 13 of pregnancy and the estrous cycle. Endometrial secretion of P₄ did not differ in the course of early pregnancy and on the respective days of the estrous cycle. The gravid myometrium was the highest source of P₄ in pregnant pigs on days 12 to 13. The release of P₄ by the cyclic myometrium rose during the examined days of the estrous cycle. The steroidogenic activity of the uterus, as described in this study, may support early pregnancy or the luteal phase of the estrous cycle in pigs.     

Temporal sequence of events in the estrous cycle of the bitch.

Serum concentrations of luteinizing hormone (LH), estradiol-17 beta, and progesteron were compared with the onset of diestrus in Beagle bitches. The LH peak occurred 8.0 (standard error (SE), 0.3) days before the onset of diestrus. Vaginal cytologic characteristics during proestrus were studied, and vaginal cornification was complete 12.1 (SE,0.2) days before the onset of diestrus. A time sequence of hormonal, vaginal cytologic, and developmental events was outlined during proestrus, estrus, and early diestrus.